The ability to replace defective or absent genes has attracted wide attention as a method to treat a variety of human diseases (Crystal 1995 Science 270:404), Lever et al. 1995 Gene Therapy. Pearson Professional, New York p. 1-91; Friedmann 1996 Nature Med. 2:144). Although originally intended as a means of correcting inherited disorders in certain populations of somatic cells, gene-based therapy can be a useful means to supply exogenous gene products to the circulatory system for the treatment of a wide range of systemic disorders that involve deficiencies in circulating proteins, such as hormones, growth factors, and clotting proteins (Lever et al. 1995 supra; Buckel 1996 TiPS 17:450), as well as a means of administering other polypeptide drugs. The success of this application depends upon developing effective methods to both manufacture the desired protein in vivo and then secrete it into blood (Crystal 1995 supra; Lever et al. 1995 supra).
Currently, DNA-based therapy (i.e., gene therapy) is carried out in a variety of ways but involves two general protocols. In the first method, referred to as ex vivo gene therapy, cells are extracted from an individual and subjected to genetic manipulation. After genetic material has been properly inserted into the cells, the cells are implanted back into the individual from which they were removed. Persistent, in vivo expression of the newly implanted genetic material after transplantation of the transformed cells has been successful (see Morgan et al., Science 237:1476 (1987); and Gerrard et al., Nat. Genet. 3:180 (1993)). In the second approach to DNA-based therapy, referred to as in vivo gene therapy, cells within a living organism are transformed in situ with exogenous genetic material.
Several different methods for transforming cells can be used in accordance with either the ex vivo or in vivo transfection procedures. For example, various mechanical methods can be used to deliver the genetic material, including the use of fusogenic lipid vesicles (liposomes incorporating cationic lipids such as lipofection; see Felgner et al., Proc. Nat. Acad. Sci. U.S.A. 84:7413-7417 (1987)); direct injection of DNA (Wolff, et al., Science (1990) 247:1465-1468); and pneumatic delivery of DNA-coated gold particles with a device referred to as the gene gun (Yang et al., Proc. Natl. Acad. Sci. U.S.A. 1990; 87:1568-9572). Morsy et al. reviews several of the different techniques useful in transformation of cells ex vivo or in vivo and provides citations of numerous publications in each area (Morsy et al., JAMA 270:2338-2345 (1993)).
One method of particular interest for delivery of genetic material involves use of recombinant viruses to infect cells in vivo or ex vivo. In these methods, a virus containing the desired genetic material is allowed to infect target cells within the subject. Upon infection, the virus injects its genetic material into the target cells. The genetic material is then expressed within the target cell, providing for expression of the desired genetic material. However, it would be preferable to avoid introduction of the desired genetic material by viral infection for a number of reasons. For example, viral infection results in delivery of viral DNA in addition to the desired genetic material, which may in turn result in undesirable cellular effects such as, adverse immune reactions, productive viral replication, and adverse integration events.
There is a need in the field for a method for delivery of genetic material into a cell in vivo to provide for expression of the introduced polynucleotide and secretion of the gene product it encodes into the bloodstream. The present invention addresses this problem.